1. Field of the Invention
The present invention generally relates to devices and methods for rapidly identifying chemicals with biological activity in liquid samples, particularly automated screening of low volume samples for new medicines, agrochemicals, or cosmetics.
2. Description of the Related Art
Drug discovery is a highly time dependent and critical process in which significant improvements in methodology can dramatically improve the pace at which a useful chemical becomes a validated lead, and ultimately forms the basis for the development of a drug. In many cases the eventual value of a useful drug is set by the timing of its arrival into the market place, and the length of time the drug enjoys as an exclusive treatment for a specific ailment.
A major challenge for pharmaceutical companies is to improve the speed and efficiency of this process while at the same time maintaining costs to an absolute minimum. One solution to this problem has been to develop high throughput screening systems that enable the rapid analysis of many thousands of chemical compounds per 24 hours. To reduce the otherwise prohibitive costs of screening such large numbers of compounds, typically these systems use miniaturized assay systems that dramatically reduce reagent costs, and improve productivity. To efficiently handle large numbers of miniaturized assays it is necessary to implement automatic robotically controlled analysis systems that can provide reliable reagent addition and manipulations. Preferably these systems and the invention herein are capable of interacting in a coordinated fashion with other systems sub-components, such as a central compound store to enable rapid and efficient processing of samples.
Miniaturized high throughput screening systems require robust, reliable and reproducible methods of analysis that are sensitive enough to work with small sample sizes. While there are a large number of potential analysis methods that can successfully be used in macroscopic analysis, many of these procedures are not easily miniaturizable, or lack sufficient sensitivity when miniaturized. This is typically true because absolute signal intensity from a given sample decreases as a function of the size of the sample, whereas background optical or detector noise remains more or less constant for large or small samples. Preferred assays for miniaturized high throughput screening assays have high signal to noise ratios for very small sample sizes.
Fluorescence based measurements have high sensitivity and perform well with small samples, where factors such as inner filtering of excitation and emission light are reduced. Fluorescence based measurements therefore exhibit good signal to noise ratios even with small sample sizes. A particularly preferred method of using fluorescence based signal detection is to generate a fluorescent (emission) signal that simultaneously changes at two or more wavelengths. A ratio can be calculated based on the emission light intensity at the first wavelength divided by the emitted light intensity at a second wavelength. This ratiometric measurement of a fluorescent assay has several important advantages over other non-ratiometric types of analysis. Firstly, the ratio is largely independent of the actual concentration of the fluorescent dye that is emitting fluorescence. Secondly, the ratio is largely independent of the intensity of light with which the fluorescent compound is being excited. Thirdly, the ratio is largely independent of changes in the sensitivity of the detector, provided that is that these changes are the same for the detection efficiency at both wavelengths. This combination of advantages makes fluorescence based ratiometric assays highly attractive for high throughput screening systems, where day to day, and, assay to assay reproducibility are important.
Traditionally, there are two general ways to read fluorescence from a multi-well plate. In one arrangement, a read head is moved from well to well and at each well, there is a dwell time during which the fluorescence signal is digitized and stored into memory. Optically, this scheme is the simplest. There is only one optical assembly, one set of filters and one detector. However, depending on how many wells there are in the plate, the read time can be unacceptably long. It is not just the dwell times that contribute to the total read time. Every time the read head is moved from well to well, it takes time to accelerate and decelerate the stage used in moving the setup.
In the other arrangement, some sort of parallelism is employed. Either a picture is taken of the plate, using a CCD camera or some other imaging arrangement, or multiple optical read heads are employed. The advantage of this arrangement is a significant reduction in the read time. However, a new difficulty is introduced, namely that of normalization. When several wells are read at the same time by several read heads, the question that arises is how to make sure that these heads behave in the same way in terms of collection efficiency, detector sensitivity, filter quality and the like. In the case of a CCD camera, the analogous issue is one of flat-fielding. Accordingly, improved methods and systems for rapidly and accurately measuring fluorescence signals in high throughput screening environments are needed.
A multiwell plate scanner comprises a detector which is scanned continuously over wells of a multiwell plate. The scanner may also be used for scanning microarrays, bio-chips and areas of samples not having physical separations.
In one embodiment, the invention is directed to a method of detecting light emitting molecules in wells of a multiwell plate. The method comprises positioning a light collector to one side of a first well of the multiwell plate continuously moving the light collector relative to the multiwell plate such that the light collector passes a first edge of the first well, passes over the first well, and passes a second edge of the first well. Fluorescent light intensity is measured during at least a portion of the time the light collector is over the first well.
The scanner may be used in a high throughput screening system comprising a storage and retrieval module, a sample distribution module, a reagent distribution module, and a detector which incorporates the scanner. One embodiment of the invention thus comprises a high throughput drug discovery method comprising retrieving chemicals from a chemical storage and retrieval module, placing the chemicals into wells of multi-well plates, scanning the multi-well plates in a substantially continuous raster scan pattern so as to detect a chemical or biological activity of one or more of the chemicals. Alternatively, the scan pattern could be in a spiral, concentric circle, or any other suitable mathematical function, depending on the shape of the sample or sample container.
The present invention is also directed to compositions and therapeutics identified by the disclosed methods. One such embodiment comprises a medicament made by a process comprising identifying a pharmacologically active chemical by a process comprising retrieving chemicals from a chemical storage and retrieval module, placing the chemicals into wells of multi-well plates, and scanning the multi-well plates in a substantially continuous plowman""s fashion so as to detect a pharmacological activity of one or more of the chemicals. Following identification, an effective amount of at least one of the pharmacologically active chemicals is incorporated into a biocompatible carrier.
A further aspect of the present invention is a method of testing a therapeutic for therapeutic activity and toxicology by identifying a compound using a method of the present invention and monitoring the toxicology and efficacy of the therapeutic in an in vivo model.